Cell-to-cell interactions are essential for the formation of dental plaque. A continuous layer of Streptococcus sanguis SA-1 cells fixed to a solid surface has been used to evaluate interactions among this bacterium, Haemophilus parainfluenzae, and Streptococcus sobrinus. S. sanguis cells were attached to a Falcon 3001 tissue culture plates or bovine enamel chips, coated with a biological adhesive. Scanning electron microscopy of the chips showed the streptococci as a contiguous surface. Radiolabeled bacteria were used to measure a second-species interbacterial adherence to the streptococcal-coated culture plates. Strains of H. parainfluenzae known to coaggregate (strain HP-28) and not to coaggregate (strains HP-42 and HP-80), in suspension with S. sanguis strain SA-1, were studied for adherence. Ten-fold-higher numbers of coaggregating strain HP-28 adhered in vitro to the streptococcal layer than did the non-coaggregating strains. S. sobrinus strain 6715 did not show appreciable adherence to the S. sanguis surface. Saliva did not affect the adherence of coaggregating or non-coaggregating H. parainfluenzae strains to S. sanguis strain SA-1. Bovine enamel chips, coated with streptococci, mounted on modified orthodontic appliances and placed in the mouths of three volunteers, facilitated the measurement of interbacterial adherence in vivo of streptomycin-resistant strains of H. parainfluenzae (HP-28R or HP-42R). Suspensions of bacteria were placed into the mouth, distributed throughout, and expectorated. After 15 or 120 minutes, the appliance with the chips was removed, the chips sonified, and colony-forming units (CFU) of streptomycin-resistant haemophili determined per chip. Similar to the in vitro experiments, ten times as many coaggregating HP-28R than non-coaggregating HP-42R adhered in vivo to the S. sanguis SA-1 layer in two of the three subjects. The measurement per mm 2 in vitro and in vivo of cell-to-cell interactions facilitates accurate comparisons between the different strains and species and will facilitate studies on the interbacterial interactions involved in plaque formation.
T he periodontal status of three groups of women; pregnant, taking oral contraceptives, and nonpregnant, were evaluated clinically and microbiologically for changes in their gingiva and any corresponding changes in the subgingival microbial plaque, specifically the percentage of Fusobacterium species and Bacteroides species. Overall, the women had relatively good gingival health. However, statistically significant increased scores were observed in the Gingival Index and the gingival crevicular fluid flow in the pregnant group compared with the nonpregnant group. The most dramatic microbial changes were the increased proportions of Bacteroides species both in the pregnant group and the group taking oral contraceptives over the nonpregnant group. Increased female sex hormones substituting for the napthaquinone requirement of certain Bacteroides were most likely responsible for this increase. No statistically significant clinical difference was noted between the group taking oral contraceptives and the nonpregnant group, although a 16‐fold increase in Bacteroides species was observed in the group taking oral contraceptives.
Developing dental bacterial plaques formed in vivo on enamel surfaces were examined in specimens from 18 adult volunteers during the first day of plaque formation. An intraoral model placing enamel pieces onto teeth was used to study bacterial plaque populations developing naturally to various cell densities per square millimeter of surface area of the enamel (W. F. Liljemark, C. G. Bloomquist, C. L. Bandt, B. L. Philstrom, J. E. Hinrichs, and L. F. Wolff, Oral Microbiol. Immunol. 8:5-15, 1993). Radiolabeled nucleoside incorporation was used to measure DNA synthesis concurrent with the taking of standard viable cell counts of the plaque samples. Results showed that in vivo plaque formation began with the rapid adherence of bacteria until ca. 12 to 32% of the enamel's salivary pellicle was saturated (ca. 2.5 x 10(5) to 6.3 x 10(5) cells per mm2). The pioneer adherent species were predominantly those of the "sanguis streptococci." At the above-noted density, the bacteria present on the salivary pellicle incorporated low levels of radiolabeled nucleoside per viable cell. As bacterial numbers reached densities between 8.0 x 10(5) and 2.0 x 10(6) cells per mm2, there was a small increase in the incorporation of radiolabeled nucleosides per cell. At 2.5 x 10(6) to 4.0 x 10(6) cells per mm2 of enamel surface, there was a marked increase in the incorporation of radiolabeled nucleosides per cell which appeared to be cell-density dependent. The predominant species group in developing dental plaque films during density-dependent growth was the sanguis streptococci; however, most other species present showed similar patterns of increased DNA synthesis as the density noted above approached 2.5 x 10(6) to 4.0 x 10(6) cells per mm2.
Fourteen freshly isolated strains of Streptococcus sanguis were obtained from the dental plaque of five healthy adults. Whole saliva was collected concomitant with the plaque isolates from the five subjects, and a second whole saliva sample was collected 10 weeks later. All possible combinations of the first five saliva samples, the second five saliva samples, and 14 strains of bacteria were tested for aggregation. Of the 140 combinations examined, 108 of 140 (77%) of the strains aggregated with the first saliva samples and 95 of 140 (68%) aggregated with the second saliva samples. Overall, 72% of the strains aggregated with both the first and second saliva samples. Removal of immunoglobulin A (IgA) from these same salivas resulted in 38 of 108 (35%) of the aggregates decreasing in intensity with the first saliva samples and 27 of 95 (29%) of the aggregates decreasing in intensity with the second saliva samples. No aggregates increased in intensity with saliva samples when IgA had been removed. Removal of IgA from saliva also resulted in a mean decrease of 46% in adherence of S. sanguis to hydroxyapatite coated with the IgA-deficient saliva. Several strains of S. sanguis were shown to aggregate strongly with human salivary and colostral IgA. In addition, S. sanguis strain S7 showed a 31% stimulation of adherence to hydroxyapatite precoated with human salivary IgA over the uncoated controls. Stepwise removal of IgA from saliva resulted in a decrease in aggregation intensity from strong (4+) to weak (1+ to 2+). Similarly, the adherence of S. sanguis to hydroxyapatite coated with these saliva samples decreased linearly as the salivary IgA was depleted. Alternatively, the addition of a small quantity of salivary IgA (20 μg/ml) to progressively diluted saliva maintained a high level of adherence and strong aggregation until the saliva dilutions reached between 1:8 in the adherence experiments and 1:32 for the aggregations. These data indicate that salivary IgA may play an important role in the microbial ecology of human dental plaque formation.
Recoveries of Streptococcus mutans from human dental plaque were lower when plated on mitis-salivarius agar obtained from Baltimore Biological Laboratories as compared with mitis-salivarius agar obtained from Difco Laboratories. However, no difference in recoveries of established laboratory strains of S. mutans was observed between these two agar preparations.
Several compounds were evaluated in an in vitro assay system for their ability to block the adherence of Streptococcus sanguis to saliva-coated hydroxyapatite and Streptococcus mutans to dextran-coated hydroxyapatite. Fatty acids, ranging from C-12 to C-20, the enzyme amylase, chlorhexidine, human sera, and several serum proteins blocked S sanguis adherence to saliva-coated hydroxyapatite. Chlorhexidine blocked S mutans adherence to dextran-coated hydroxyapatite, but human sera and serum proteins did not. The effects of these compounds on the adherence of these organisms to hydroxyapatite may help in the development of specific plaque control methods for use in human populations.
